摘要
目的建立大鼠血浆中齐墩果酸的LC-MS/MS测定法,并应用于灌胃白花蛇舌草提取物齐墩果酸在大鼠体内的药动学研究。方法取大鼠血浆100μL,加入10μL内标甘草次酸(5μg·mL-1),依次用乙酸乙酯800μL提取两次,上清液用氮气流吹干,100μL流动相复溶,高速离心10 min,取上清液5μL进行LC-MS/MS测定。色谱条件:Agilent 1200高效液相色谱系统,色谱柱Phenomenex Gemini 110A C18色谱柱(50 mm×2.0mm,5μm);流动相:乙腈-0.01%甲酸水梯度洗脱;柱温为40℃;流速为0.3 mL·min-1。质谱:API4000负离子MRM模式,齐墩果酸和甘草次酸的选择检测离子质荷比分别为m/z 455.3→455.3,m/z 469.4→425.3。结果齐墩果酸血浆浓度线性范围0.5～250 ng·mL-1,线性关系良好(r=0.9979),最低检测限(LOQ)为0.1 ng·mL-1,提取回收率为62.76%～65.38%,日内精密度RSD为3.55%～7.45%,日间精密度RSD为2.34%～7.60%,稳定性均符合方法学测定要求,血浆无内源性基质干扰。结论该法简便,灵敏,重复性好,可应用于测定血浆中齐墩果酸的含量并应用于灌胃白花蛇舌草提取物大鼠体内齐墩果酸的药物动力学研究。
Objective To establish a method for determining the plasma concentration of oleanolic acid in rat with LC-MS/MS, and to calculate the pharmacokinetie parameters of oleanolic acid in rat after oral administration of Her- ba Hedyotis Diffusae extract. Methods The internal standard glycyrrhetinic acid was added to 100 μL of rat plasma, following by liquid-liquid extraction for twice with 800 μL of ethyl acetate. After centrifugation for 10 min, the su- pernatant were dried by nitrogen and recombinanted by 100 μL mobile phase, and 5 μL of supernatant was used for LC-MS/MS. Agilent 1200 HPLC system with Phenomenex Gemini C18（50 mm × 2.0 mm, 5 μm） was used. Mo- bile phase consisted of aeetonitrile and 0.01% formic acid solution at a flow rate of 0.3 mL·min^-1. The temperature of column was 40 ℃. The LC-MS/MS system was operated by using electrospray ionization probe in the negative ion mode; Scan mode was in multiple reaction ion monitoring（MRM） mode. The ion of monitor was m/z 455.3→455.3 for oleanolic acid, m/z 469.4→425.3 for glycyrrhetinic acid （internal standard）, respectively. Results The linear ranges of oleanolic acid was from 0.5 to 250 ng·mL^-1 （r=0.9979）. The low limit of quantification （LOQ） of oleanolic acid was 0.1 ng·mL^-1. The extraction recovery of oleanolic acid was from 62.76 % to 65.38 %. The RSD of intra-and inter-day variation ranged from 3.55 % to 7.45 %, and 2.34 % to 7.60 %, respectively. The method had good sta- bility, without any endogeneous interference. Conclusion A sensitive, accurate and suitable LC-MS/MS method for determining oleanolic acid has been developed and successfully applied to the pharmacokinetic study of Herba Hedy- otis Diffusae extract after oral administration in rats.
出处
《中药新药与临床药理》
CAS
CSCD
北大核心
2012年第4期441-445,共5页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
2011国家自然基金资助项目(81102883)
广东省自然科学基金资助项(S2011010005540)
广东省优秀博士学位论文基金(粤学位函(2010)1号)
科技创新重点项目(粤科教函(2011)83号)
